Primary biliary cirrhosis (PBC) is an autoimmune disease of the liver characterized by immune mediated destruction of bile ducts and high titer autoantibodies to mitochondrial components. The mitochondrial autoantigens are members of the 2-oxo acid dehydrogenase enzyme complex (the major autoantigen being PDC-E2). We have mapped B. and T cell autoepitopes in PDC-E2 and demonstrated heterogeneous TCR usage in PDC-E2 specific T cells. Further, we have made the important observation that there are markedly increased amounts of a molecule with some, but not all, of the properties of PDC-E2 on biliary epithelial cells (BEC) of PBC but not control patients. This is the only known difference between the BEC of PBC patients and controls and provides a clue as to why immune damage is focused on biliary epithelium. We have thus developed a hypothesis, we propose to test, that explains the selective destruction of BEC and the mechanisms involved in the immunopathology of PBC. Firstly, we will identify the molecule uniquely expressed in BEC in PBC. We will define whether the antigen is a cross-reactive molecule that shares a single and/or multiple autoantigenic epitopes, a truncated or otherwise processed form of PDC-E2, or a tissue specific isoform. If this is a cross-reactive molecule we will clone and sequence the gene encoding it. Secondly, we will determine if the increased expression of antigens in the BEC of PBC is limited to a PDC-E2 like molecule or whether there is also increased expression of molecules with similarity or crossreactivity to the other commonly recognized autoantigens BCOADC-E2 and OGDC-E2. The answer to this question will differentiate between widespread molecular mimicry or global upregulation of endogenous sequences. Finally, we will determine the temporal changes in the specificity of the immune response in PBC. We will determine whether the PDC-E2 like molecule on BEC is found in asymptomatic and early stage PBC and follow its expression during disease progression. We will also determine if there is a temporal progression in the cellular response in PBC and, if so, whether the peptides that induce the T cell responses can be identified and the precise MHC antigens that present such peptides be defined. Similarly, we will address the issues of progression of specific TCR V/beta and peptide-binding motifs. The availability of cDNAs for the autoantigens, cDNA BEC libraries, murine mAbs, combinatorial Abs, and an extensive network of collaborators/tissue availability places our laboratory in a unique position to address fundamental mechanisms that will advance our understanding of both PBC and autoimmunity in general.